Classification

Higher order taxa

Species

Strains

Laboratory: NCFM, 4962, CNRZ216, CNRZ218

Human:HA1, HA2, HA3, HM2, HM6

Pig: PA3, PA12, PA19, P18, P47

Chicken: C1, C2, C3, C7, C11

Description and Significance

In general, Lactobacilli is the largest genus of the lactic acid bacteria group and includes over 50 species. Lactobacilli commonly inhabit the gastrointestinal (GI) tract, oral, and vaginal regions of humans and animals.

Lactobacilli have many important roles in industry. They contribute to the production of some cheeses, yogurt, and other products. The lactic acid produced by Lactobacilli inhibits the growth of other organisms and lowers the pH of the product in these products. The starter cultures for such products are carefully cultivated and maintained because their metabolic end products contribute to the flavor of the final food product. Additionally, some of Lactobacilli's metabolic reactions are intentionally manipulated to breakdown milk proteins during cheese production.

Early studies of L. acidophilus were performed on strains isolated from fecal material of humans, pigs and chickens. Since then L. acidophilus has been further characterized as a short Gram-positive rod (2-10μm), is homofermentative and has optimal growth at temperatures of 37˚C-42˚C. Of the Lactobacillus species, L. acidophilus is the most well known and is commercially distributed as a probiotic. The World Health Organization defines a probiotic as "live microorganisms which, when administered in adequate amounts, confer a health benefit on the host". (see Ecology). (1)

Further isolation and investigation into the physiological, biochemical, genetic, and fermentative properties have been widely explored in both humans and animals. The L. acidophilus strain, NCFM, was isolated from a human in 1970 and characterized at North Carolina State University. NCFM has been commercially available in the United States as a probiotic strain since the mid-1970s. NCFM is also used for formula, yogurt and fluid milk production. (2)

Genome structure

The complete circular genome of the NCFM strain of L. acidophilus contains 1,993,564 nucleotides. The DNA GC content was determined to be 34.71%. There are 1,864 open reading frames (ORFs) and 72.5% have been classified functionally. (2)

L. acidophilus NCFM contains no plasmids. No complete prophages were found, but 9 phage-related integrases are predicted to remain as remnants. Additionally, 3 unique regions classified as potential autonomous units (PAUs) were identified. PAUs have unique characteristics that suggest that they are phage or plasmid remnants, but have not be further classified at this point. A unique signature sequence was noted. It is composed of 32 repeats of 29 base pairs (bps). (2)

L. acidophilus NCFM produces a bacteriocin, called lactacin B that demonstrates antimicrobial activity against other Lactobacilli and Enterococcus faecalis. The molecular mass of lactacin B is 6500 Daltons and is sensitive to proteinase K and pronase. It is stable between temperatures ranging from 121˚C to -20˚C. (3)

Proteins that allow for L. acidophilus's unique metabolic functions and ability to survive acid conditions were also identified. These include gene clusters that allow for transport of a diverse group of carbohydrates, such as fructooligosaccharides and raffinose, and were often associated with transcriptional regulators. Coupled regulatory systems between acid tolerance and bacteriocin production are predicted in 9 locations. Overall these genes enable L. acidophilus to survive in the harsh GI conditions and promote interaction with the intestinal mucosa and microbiota. (3)

Cell structure and metabolism

In 1999, an H+ induced ATPase was identified in L. acidophilus. Based on primary structure and the genetic organization, it was further classified as a F1F0-type ATPase. Its similarity to the streptococcal ATPase and the H+ inducibility of the operon suggests that it is responsible for an ATP-dependent exclusion of protons in order to maintain cytoplasmic pH (~7). (4)

L. acidophilus lack cytochromes, porphyrins, and respiratory enzymes and as a result are unable to undergo any oxidative phosphorylation or respiration. Because they utilize sugars as their substrates for fermentation, they inhabit environments with high sugar abundance, such as the GI tract in humans and animals. More specifially, L. acidophilus is homofermentative which means that the only byproduct it forms from fermentation is lactic acid. For every one glucose molecule that undergoes fermentation in L. acidophilus, the energy yield is two ATPs. As a result, homofermentative microbes must catabolize large amounts of substrate to generate enough energy for growth. In addition to glucose, L. acidophilus utilizes aesculin, cellobiose, galactose, lactose, maltose, salicin, sucrose, and trehalose for fermentation.

Ecology

L. acidophilus is best known as a probiotic. The exact mechanism of the probiotic effect is still under investigation. (see Current Research)

Adherence and colonization is one of many suggested mechanisms responsible for the probiotic effect of L. acidophilus. Adherence and colonization of the intestinal epithelium can act in two ways: (1) competition for space on the epithelium and (2) interaction with enterocytes. There is some evidence that L. acidophilus NCFM has the ability to adhere through a protein mediated mechanism. (see Current Research) After L. acidophilus has colonized the GI tract, as a probiotic bacteria it has the potential to influence the existing microbial population in favor of the host's health. (3)

Antimicrobial activity is considered an important mechanism by which probiotic bacteria act to inhibit a range of microbes that have potentially detrimental effects. It is suggested that L. acidophilus produces bacteriocins (proteins that are active against other bacteria). This specific mechanism is currently being researched (see Current Research). (3)

L. acidophilus has been suggested as a supplement for lactose intolerant individuals. When taken orally and in sufficient dosages, there is evidence for a decrease in symptoms of lactose maldigestion. Presumably, the L. acidophilus colonizes the GI tract and contributes to the metabolism of lactose during digestion and transit through the GI tract. (3)

Current Research

Protein Mediated Adherence

Adherence to the epithelium of the GI tract is thought to play an important role in the probiotic effect of L. acidophilus. Stuies show that the exact mechanism of adherence varies from strain to strain. Possible mechanisms include protein and carbohydrate mediated adherence. Although both types of mechanisms have been demonstrated in vitro neither has successfully been demonstrated in vivo. Because the GI tract is a constantly changing environment it is difficult to mimic the environment in vitro. Further studies with biopsies of intestinal tissues are necessary to confirm adherence and retention of L. acidophilus in the GI tract. However, studies that include such biopsies are rare. In vitro, NCFM specifically shows a protein mediated response. The NCFM that demonstrated adherence did not appear to have a polysaccharide layer, which may be significant to its ability to adhere. Further study is needed to confirm mechanisms in vivo. (3)

Innate Antimicrobial Activity

When L. acidophilus is co-cultivated with other organisms, L. acidophilus has repeatedly been shown to inhibit the growth of competing microbes. It is thought that L. acidophilus produces a variety of antimicrobial compounds including organic acids, hydrogen peroxide, diacetyl and bacteriocins. The activity of these compounds is evident in the laboratory, but the in vivo role of these compounds is less clear. This is an area of active research. For instance, human fecal samples show a correlation between a reduction in pH and an increase in short chain fatty acids with higher fecal counts of Lactobacilli and bifidobacteria (which is another species that exhibits a probiotic effect). In the laboratory strain NCFM demonstrated antagonistic activity against common foodborne disease agents such as Staphylcoccus aureus, Salmonella typhimurium, and enteropathogenis Escherichia coli. (3)

Probiotic Effects Beyond the GI Tract

L. acidophilus has long been considered a probiotic conferring digestive benefits to its host. Now research is beginning to examine L. acidophilus' potential probiotic effect in combating vulvovaginal candidiasis (VVC). VVC is associated with a low number of Lactobacilli in the vagina or with peroxide-non-producing vaginal Lactobacilli. A review of the literature shows no clear indication of whether or not L. acidophilus is effective in all VVC cases. The studies are preliminary and had methodological problems such as small sample size, lacked a control group and included women without confirmed VVC. Further studies are necessary with appropriate experimental design. Additionally, studies regarding oral or intravaginal dosage are necessary to determine a more efficient administration of the probiotics. (5)